Progress in the treatment of solid tumors has been slow and sporadic despite the development of new chemotherapeutic agents. There are many roadblocks to successful chemotherapy, including drug resistance, resistance to apoptosis, and the inactivation of tumor suppressor genes. Some human cancers are drug resistant before treatment begins, while in others drug resistance develops over successive rounds of chemotherapy.
One type of drug resistance, called multidrug resistance, is characterized by cross resistance to functionally and structurally unrelated drugs. Typical drugs that are affected by the multidrug resistance are doxorubicin, vincristine, vinblastine, colchicine, actinomycin D, and others. At least some multidrug resistance is a complex phenotype that is linked to a high expression of a cell membrane drug efflux transporter called Mdr1 protein, also known as P-glycoprotein. This membrane xe2x80x9cpumpxe2x80x9d has broad specificity and acts to remove from the cell a wide variety of chemically unrelated toxins.
Another factor in cancer therapy is the susceptibility of targeted cells to apoptosis. Many cytotoxic drugs that kill cells by crippling cellular metabolism at high concentration can trigger apoptosis in susceptible cells at much lower concentration. Increased susceptibility to apoptosis can be acquired by tumor cells as a byproduct of the genetic changes responsible for malignant transformation, but most tumors tend to acquire other genetic lesions which abrogate this increased sensitivity. Either at presentation or after therapeutic attempts, the tumor cells can become less sensitive to apoptosis than vital normal dividing cells. Such tumors are generally not curable by conventional chemotherapeutic approaches. Although decreased drug uptake, altered intracellular drug localization, accelerated detoxification and alteration of drug target are important factors, pleiotropic resistance due to defective apoptotic response is also a significant category of drug resistance in cancer.
An important tumor suppressor gene is the gene encoding the cellular protein, p53, which is a 53 kD nuclear phosphoprotein that controls cell proliferation. Mutations to the p53 gene and allele loss on chromosome 17p, where this gene is located, are among the most frequent alterations identified in human malignancies. The p53 protein is highly conserved through evolution and is expressed in most normal tissues. Wild-type p53 has been shown to be involved in control of the cell cycle, transcriptional regulation, DNA replication, and induction of apoptosis.
Various mutant p53 alleles are known in which a single base substitution results in the synthesis of proteins that have quite different growth regulatory properties and, ultimately, lead to malignancies. In fact, the p53 gene has been found to be the most frequently mutated gene in common human cancers, and is inactivated in xcx9c50% of tumors. It is particularly associated with those cancers linked to cigarette smoke. The overexpression of p53 in breast tumors has also been documented.
The increase in p53 protein in response to various stresses is an important regulator of cell cycle and apoptosis. In transcription-dependent p53 activation, p53 functions as a site-specific transcription factor that induces p53-inducible genes such as p21cip1/waf1, bax, gadd45, and mdm2. This in turn initiates the program of growth arrest or apoptosis in a stress-specific and/or cell type-specific manner. Two recent studies have shown that p53-mediated induction of p21 inhibits the apoptotic response by inducing growth arrest. For example, p53- or p21-deficient cells have been shown to be more sensitive to adriamycin (doxorubicin)-induced apoptosis than wild-type cells because they do not induce p21 in response to doxorubicin. Therefore, a compound that blocks chemotherapy-mediated growth arrest may accelerate and enhance apoptosis.
An area to search for new therapeutic interventions is that of traditional Chinese medicines. One of these traditional medicines is from Tripteryguim wilfordii Hook F, a shrub-like vine from the Celastraceae family. A variety of preparations derived from this plant have been used in South China for many years to treat different forms of arthritis and other autoimmune diseases. In 1978, an extract of Tripterygium wilfordii Hook F was produced by chloroform methanol extraction of the woody portion of the roots and designated T2. Reports in the Chinese literature describe T2 treatment of more than 750 patients with a variety of autoimmune diseases.
The Chinese experience has suggested that a daily dosage of about 1 mg/kg of T2 is safe and effective as an immunosuppressant. Acute and chronic toxicity studies have been carried out in China using a variety of animal models. The LD50 in mice was reported to be around 150 mg/kg. The toxicity studies suggest that T2 exhibits a reasonable safety index and should be able to be administered to patients safely.
The development of chemotherapeutic agents and combinations of agents that avoid problems of drug resistance and resistance to apoptosis are of great interest for the treatment of cancer.
Literature
The isolation, purification, and characterization of immunosuppressive compounds from tripterygium: triptolide and tripdiolide is reported by Gu et al. (1995) Int J Immunopharmacol 17(5):351-6. Yang et al. (1998) Immunopharmacology 40(2):139-49 provide evidence that suggests the immunosuppressive agent triptolide inhibits antigen or mitogen-induced T cell proliferation, and induces apoptotic death of T cell hybridomas and peripheral T cells. Shamon et al. (1997) Cancer Lett 112(1):113-7 evaluate the antitumor potential of triptolide. Tengchaisri et al. (1998). Cancer Lett. 133(2):169-75 evaluate the antitumor activity of triptolide against cholangiocarcinoma growth in vitro and in hamsters.
Lee et al. (1999) J Biol Chem 274(19):13451-5 describe the interaction of PG490 (triptolide) with tumor necrosis factor-alpha to induce apoptosis in tumor cells. Triptolide was found to inhibit T-cell interleukin-2 expression at the level of purine-box/nuclear factor of activated T-cells and NF-kappa B transcriptional activation by Qiu et al. (1999) J Biol Chem. 274(19):13443-50.
Chang et al. (2000) J. Biol. Chem. 276:2221-2227 describes the cooperation of triptolide with chemotherapeutic agents in tumor cell apoptosis.
Compositions and methods are provided for the use of diterpenoid triepoxides in combination with anti-proliferative agents, as a combination therapy to treat hyperproliferative disorders. The methods and compositions are particularly useful in the treatment of multi-drug resistant tumor cells. Anti-proliferative agents of interest include agents active in killing tumor cells, as well as immunosuppressants, and a variety of other agents that reduce cellular proliferation in targeted tissues. The targeted cells are contacted with an anti-proliferative agent and diterpenoid triepoxides, e.g. triptolide, tripdiolide, etc., or prodrugs that convert to such compounds under physiological conditions, either locally or systemically. Synergistic combinations provide for comparable or improved therapeutic effects, while lowering adverse side effects.
The combination therapy is particularly effective in the treatment of tumor cells that express p21waf1/cip1, which cells can otherwise be resistant to the induction of apoptosis. In one embodiment of the invention, tumors are assayed for the expression of p21 to determine whether they will benefit from the combined therapy of the present invention.